An understanding of the ways in which microbial toxins and viruses act at the cellular and molecular level will enable better ways of controlling or abrogating their negative effects on cells and tissues to be designed. The purpose of this project is to isolate and study mutant cultured cells, or human and animal origin, in order to investigate mechanisms of Pseudomonas exotoxin A action, replication of animal viruses, receptor-mediated endocytosis, and the cellular pathways and mechanisms involved. Ongoing studies of a group of cell strains already isolated, that are resistant to Pseudomonas toxin and certain viruses because of diverse genetic lesions will be continued. New strains will also be isolated and characterized, using biological, biochemical, immunological, and genetic techniques. Ligands with known or postulated mechanisms of entry, such as well-characterized viruses, lectins, and selected hormones and transport proteins, will be used as probes to determine the biochemical and genetic basis of resistance in the mutant cells. Monoclonal antibodies to the Pseudomonas toxin receptor will be prepared, in order to isolate, purify, and quantify the receptor, to study its structure-function relationships, and to use as probes for cloning the toxin receptor gene. Monoclonal antibodies to Sindbis virus glycoproteins will be prepared for the purpose of localizing, within the cell, the site of an endopeptidase activity essential for the processing of these glycoproteins, and, it is postulated, for the activation of Pseudomonas toxin. The gene (or genes) for this enzyme activity will be cloned, taking advantage of a group of resistance mutant cells that are deficient in this activity. A unique opportunity to investigate not only sensitivity and resistance to microbial toxins, but also regulation of gene expression and cellular function, and viral replication, is presented.